1. Field of the Invention
The present invention relates to a method for manufacturing red mold dioscorea, and more particularly relates to a method for manufacturing red mold dioscorea with a solid state cultivation method or a liquid state cultivation method.
2. Description of the Prior Art
Hypertension caused by hypercholesterol is a popular civilization disease in recent years. Following the advancement of medical science, populations of middle-aged and geriatric people are increasing. It is difficult to eliminate superoxide free radicals in middle-aged and geriatric people result from the lower metabolic mechanism, and the middle-aged and geriatric diseases include cancer and cardiopathy are prone to occur. Cardiovascular diseases arise from hypercholesterolemia include apoplexy, coronary atherosclerotic cardiopathy and hypertension. In pursue of the healthy diet in life, health food nowadays is very popular. Red mold products possess both effects of inhibition of cholesterol synthesis and blood pressure lowering and become one of the popular health food.
Numerous researches in recent years prove that red mold rice have significant lipid-lowering effect on hyperlipidemia hamsters, thus red mold rice have potential to be an ingredient in cholesterol-lowering drugs and improve this kind of disease. The above effects are result from that red mold rice can produce several kinds of secondary metabolite, such as cholesterol-synthesis inhibitor, blood pressure lowering material, anti-cancer material and so forth. Some previous researches about secondary metabolite of red mold yeast indicate that the yield and pattern of secondary metabolite may be directly or indirectly influenced by cultural environment and method.
Recently, red mold related products made by using some specific fermented substrate are proposed. In 2006, Lee et al. propose their study of “Monascus fermentation of dioscorea for increasing the product of cholesterol-lowing agent”, and the study is then published on Journal of Applied Microbial and Cell Physiology. In Lee's study, Monascus-fermented substrate, for example, Monascus fermented dioscorea, is proven its effect on increasing the production of monaclin K and monascin, wherein the monaclin K can be used as a cholesterol-lowering agent and the monascin can be as a anti-inflammation agent.
Please refer to FIG. 1, there is shown a flowchart of a red mold substrate preparation method proposed by Lee. As shown in FIG. 1, to prepare the red mold substrate, a substrate is firstly purchased from local supermarket (S01′). Then, five grams of substrate is soaked in distilled water for 8 hr (S02′), and excess water is removed by using a sieve (S03′). After the steps of S01′˜S03′ are finished, the substrate is then autoclaved in a wood dish at 120° C. for 20 min (S04′). And after being cooled, the substrate is inoculated with a 5% spore suspension (107 spores/ml) and 0.3% ethanol (S05′). Therefore, the inoculated substrate is cultivated at 30° C. for 10 days (S06′); moreover, during the culturing stage, 145 ml of water is added once every 12 hr at a total of three times and the addition of water starts on the fifth day of cultivation. Therefore, Lee successfully stimulates the formation of monacolin K by adopting above-mentioned preparation method. In addition, Lee also finds that the monascus-fermented dioscorea have more amount of yellow pigment than monascus-fermented rice. Yellow pigment of Monascus species including monascin and ankaflavin has been shown as an anticancer agent and anti-inflammation agent.
Therefore, because Lee's preparation method is proven the effect on increasing the production of monaclin K and monascin, the preparation method is then widely used for the production of monaclin K and monascin, and called solid state cultivation method. However, in spite of Lee's preparation method is helpful to the production of monaclin K and monascin, his preparation method still includes the shortcomings and drawbacks as follows:
1. In the steps S02′ and S03′ of Lee's preparation method, the substrate is soaked in distilled water for 8 hr and the excess water is removed by using a sieve. The person skilled in this art is able to find that Lee still fails to optimize a best water ratio for soaking the substrate, and the water ration of steps S02′ and S03′ is obviously unsuitable to the sterilization of the substrate.2. In Lee's preparation method, it does not provide a suitable environment for culturing the inoculated substrate, for example, a suitable environment with an optimal relative humidity. Water supplementation is used in Lee's preparation method, but the environment humanity control is more useful to stabilize the production of monacolin K, monascin, ankaflavin, and citrinin, the person skilled in this art knows that.3. Although Lee's preparation method is proven the effect on increasing the production of monaclin K and monascin can be increased, it also helpful to increase the production of citrinin, which is grouped into a kind of polyketide derivatives of Monascus. So that Lee adds 0.3% (v/w) ethanol to substrate (dioscorea) in order to lower citrinin production. However, the procedure also significantly decreased the production of the formational metabolites monascin and ankaflavin by 2.5 and 3-fold.
In addition, differing from the solid state cultivation method for increasing the production of monaclin K and monascin, in 2007, Lee further proposes his study of “Improving the ratio of Monacolin K to citrinin production of Monascus purpureus NTU 568 under dioscorea medium through the mediation of pH value and ethanol addition”, and the study is then published on Journal of Agricultural and Food Chemistry. In this study, Lee proposes a liquid state cultivation method for increasing the production of monaclin K and monascin. Please refer to FIG. 2, which illustrates a flowchart of the liquid state cultivation method proposed by Lee. As shown in FIG. 2, to prepare and culture the monascus-fermented dioscorea (substrate), a dry dioscorea root is firstly milled into powder (60-80 mesh) and used as the substrate of seed culture (S01″), and a culture medium with 5% the powdered dioscorea is then prepared (S02″). After the steps of S02′˜S02′ are finished, the culture medium is subsequently inoculated with 5% Monascus spore solution and 0.3% ethanol (S03″). Therefore, in step S04″, the fermentation is carried out with aeration rate at 4 vvm, 30° C. and the agitation at 200 rpm for 12 days (S04″). After fermentation, the dioscorea is collected and dried at 55° C. for 48 hr (505″), and eventually to weight the total dry biomass for the calculation of the ratio of dry biomass weight to dioscorea added weight (B/D ratio) (S06″).
Through the liquid state cultivation method, Lee successfully stimulates the formation of monacolin K and monascin. Besides, Lee also determines an optimum culture conditions based on experiment results. However, in spite of Lee's liquid state cultivation method is helpful to the production of monaclin K and monascin, his method still includes the main drawback of “lacking citrinin-inhibiting step”.
So that, in view of the above-mentioned conventional red mold substrate preparation method (including the solid state cultivation method and the liquid state cultivation method) still have shortcomings and drawbacks, it is necessary to choose a fermented substrate with better effects and adjust the culture conditions to be optimum so as to increase the production of monaclin K, monascin and ankaflavin, and then to elevate the effects of blood-lipid lowering, blood-pressure lowering and atherosclerosis prevention. Therefore, the inventor of the present invention resorted to past experience, imagination, and creativity, performed experiments and researches repeatedly, and eventually devised the present invention: a method for manufacturing red mold dioscorea.